Synthesis, Characterization and Antimicrobial Activity of Some New Benzimidazole Derivatives Bearing the Salicyl Moiety

doi:10.21203/rs.3.rs-1882010/v1

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Research Article

Synthesis, Characterization and Antimicrobial Activity of Some New Benzimidazole Derivatives Bearing the Salicyl Moiety

https://doi.org/10.21203/rs.3.rs-1882010/v1

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Sixteen new benzimidazole compounds bearing the salicyl moiety have been synthesized by cyclization of 4,5-dichloro/dimethyl-1,2-phenylenediamine with various salicylaldehydes. All the compounds were characterizated by spectral methods (FT-IR, ¹H NMR, ESI-MS and fluorescence spectroscopy) and elemental analysis. It is seen that the dichloro derivatives and 5-substitution of the phenolic ring shift the fluorescence radiation (emission) to the higher wavelength (red shift). Antimicrobial activities of the compounds were screened against six bacteria and a fungus. The dimethyl derivatives (9–16) are generally more active towards the microorganisms and show higher antifungal activity than the dichloro derivatives. The compound 13, which is effective against all microorganisms, shows the broadest activity.

Benzimidazole

antimicrobial activity

salicyl

substituent effect

Benzimidazoles nucleus, a fused heterocylic compound containing benzene and imidazole, has been studied extensively in the recent years due to its wide-range of biological properties such as antiviral [1], antimicrobial [2], anticancer [3, 4], antioxidant [5], antiallergic [6], antidepressant [7], antihelminthic [8] and lipase inhibitions [9]. Moreover, various compounds containing benzimidazol scatfold are present in many drugs such as hiabendazole, flubendazole (anthelmintic), astmizole (antihistaminic), lansoprazole and omeprazole (antiulcerative) for treatmet of numereous diseases [10].

Many benzimidazole based compounds have been found various applications in the textile industry as optical brighteners [11], or as dyes for dyeing wool and silk fibres [12]. Some of its derivatives are also used as herbicides[13], pesticides[14] and as corrosion inhibitors for metal and alloys [15–16]. Synthesis of benzimidazoles have been targets to researchers due to these various applications in great many fields.

In our previous studies, we have reported synthesis, characterization of some new biologically active benzimidazole compounds and their various metal complexes [17–21]. As a continuation to our previous work, we have synthesized sixteen new benzimidazole compounds bearing salicyl moiety (1–16, Scheme 1) in good to high yields. In vitro antibacterial and antifungal activities of these compounds were screened against selected six bacteria and a fungus. In addition, the effects of different substituents on the structural properties and antimicrobial activities of the synthesized benzimidazole derivatives were investigated.

Compound number	R₁	R₂	R₃	R₄	R₅
1 (9)	Cl (CH₃)	Cl (CH₃)	OH	H	H
2 (10)	Cl (CH₃)	Cl (CH₃)	H	OH	H
3 (11)	Cl (CH₃)	Cl (CH₃)	H	H	OH
4 (12)	Cl (CH₃)	Cl (CH₃)	H	H	OCH₃
5 (13)	Cl (CH₃)	Cl (CH₃)	H	OCH₃	H
6 (14)	Cl (CH₃)	Cl (CH₃)	OCH₃	H	H
7 (15)	Cl (CH₃)	Cl (CH₃)	H	H	CH₃
8 (16)	Cl (CH₃)	Cl (CH₃)	CH₃	H	H

The analytical data and physical properties of the compounds are summarized in Experimental Section. In this study, sixteen benzimidazolylphenol compounds containing various substituents were synthesized and their structural properties and antimicrobial activities were investigated.

The melting points of the dichloro derivatives (1–8), between 305 and 358°C, are higher than those of the dimethyl derivatives (9–16), between 276 and 324°C. It is observed that the 5-substitution raise the melting point: 5-OH derivatives, 3 and 11, have melting points of 355 and 324°C, 5-methoxy derivatives, 4 and 12, have 358 and 293°C, and 5-methyl derivatives, 7 and 15, have melting points of 349 and 285° C, respectively.

FT-IR Spectra

The FT-IR spectra of the compounds (Experimental Section) shows medium intensity absorption bands ranging between 1615–1665 cm^− 1 are ascribed to C = N stretching bands. The medium absorption bands at the 1599–1640 cm^− 1 range are related to the aromatic C = C stretching vibrations.

The characteristic ν(O–H) and ν(N–H) vibration frequencies of the compounds exhibit medium or strong bands at the 3268–3353 cm⁻¹ range in the FT-IR spectra, probably caused by doubly intramolecular hydrogen bonding between the phenoxyl hydrogen atom and one of the imine nitrogen atoms (Fig. 1).

The C–O vibrations are observed around 1220–1300 cm^− 1 as sharp bands. CH stretching bands of aromatic compounds are appeared between 3056 and 3100 cm^− 1. Characteristic out of bending of the aromatic CH {δ(C–H)} vibrations at 813–872 cm^− 1 and 729–804 cm^− 1 ranges as strong or medium bands. The C–Cl stretching vibrations of could be detected at the range 657–684 cm^− 1 as medium bands at the compounds 1–8. Methoxy (4, 5, 6, 12, 13, 14) and methyl groups (7, 8, 9–16) give medium or weak bands in the 2850–3000 cm^− 1 range due to the C–H stretch.

¹ H NMR Spectra

The ¹H NMR spectral data of the compounds is summarized in Experimental Section. The NH and OH signals appeared separately in dichloro-derived compounds (1–8) and 14, 15 and 16 (Fig. 2), which is due to weak intra-molecular hydrogen bonding whereas they are seen as a single band in 9 and 10 (12.94 and 12.90 ppm, respectively), which indicates strong intra-molecular hydrogen bonding between the imine nitrogen with double bond and phenolic hydrogen atoms (Fig. 1). In the compounds 4, 11, 12 and 13, NH and OH protons could not be observed because they were dissociated. In these compounds, NH and OH protons have a stronger acidic character than others and strong intra-molecular hydrogen bonds. The appearance of OH and NH protons as separate signals may also indicate an isomeric structure as in Fig. 3.

Fluorescence Spectra

The fluorescence spectral data are given in Experimental Section. Fluorescence spectra of the compounds were obtained in ethanol at room temperature (excitation wavelength: 354 nm; concentration: ~10⁻⁴ M).

It is seen that the dichloro derivatives (1–8) shift the emission band to higher wavelength (red-shift) according to the dimethyl derivatives. The maximum emission wavelengths of the dichloro derivatives appear at the range of 441–521 nm, while those of dimethyl derivatives (9–16) range is in 422–495 nm. It is seen that the compound 3 has the highest emission band maximum, while the compound 13 has the lowest value (Fig. 4). In addition, the 5-substitution in the phenolic ring increases the emission wavelength with respect to the others. For example, emission maximum values for 3 and 4 (5-OH and 5-OCH₃ derivatives) are 521 and 511 nm, respectively.

When the 3-, 4- and 5- derivatives are compared, it is seen that the redshift order is 5- > 4- > 3-derivatives (redshift effect, Fig. 5). For example, the emission maximum wavelength values are 500, 441 and 521 nm for 1, 2 and 3 (3 > 1 > 2). A similar effect is observed in other triad groups (4, 5 and 6; 9, 10 and 11; 12, 13 and 14 groups are compared within themselves).

Based on excited-state intramolecular proton transfer (ESIPT), the compounds show dual or triple emission. The shoulder or weak bands are attributed to the emission from a weak enol form of the compound, while the main band is attributed to the emission from tautomer form (i.e. keto form, Fig. 6) via ESIPT process [22–24]. The weak bands around 393 nm can be considered as evidence for the enol form, and medium or strong bands appearing at higher wavelength can be accepted as evidence for tautomer structure.

Antimicrobial Activity

The results of antibacterial and antifungal activities of the target compounds are presented in Table 1.

At a glance, Table 1 shows us that the dimethyl derivatives (9–16) are generally more active towards the microorganisms than the dichloro derivatives. It is observed that the compound 13 which is effective against all microorganisms, shows the broadest activity. It is seen that the compounds 7 and 11 shows the highest antibacterial activity against S. epidermidis and S. aureus, respectively, with MIC values of 2.4 µg/mL, a superior activity. Compound 11 also shows remarkable antibacterial activity against S. epidermidis (19.5 µg/mL). The fact that 11 of the compounds are effective on C. albicans led us to underline to highlight the antifungal character of the studied compounds. Another interesting observation is that the compound 8 is not effective against any microorganism.

Table 1

In vitro antimicrobial activity of the compounds (MIC, µg/mL)
Compounds	----------------------M i c r o o r g a n i s m s------------------------
	Sa^[a]	Se^[a]	Ec^[b]	Kp^[b]	Pa^[b]	Pm^[b]	Ca
1	-	-	-	78	-	-	-
2	-	-	1250	1250	-	-	156
3	78	-	312	625	625	-	19.5
4	312	-	-	625	-	-	-
5	625	-	625	1250	-	-	312
6	-	-	1250	-	-	-	312
7	78	2.4	-	-	-	-	-
8	-	-	-	-	-	-	-
9	-	-	-	-	-	-	156
10	19.5	19.5	-	-	-	-	156
11	2.4	19.5	-	-	-	-	39
12	-	-	-	-	-	-	78
13	39	39	312	156	156	312	78
14	39	156	-	-	-	625	78
15	-	-	-	-	-	-	156
16	78	156	-	1250	-	-	-
Ciprofloxacin	0.25	0.0625	0.0156	0.0078	1.0	0.0156	-
Fluconazole	-	-	-	-	-	-	1.0
Sa Staphylococcus aureus ATCC 6538 Se Staphylococcus epidermidis ATCC 12228 Ec Escherichia coli ATCC 8739 Kp Klebsiella pneumoniae ATCC 4352 Pa Pseudomonas aeruginosa ATCC 1539 Pm Proteus mirabilis ATCC 14153 Ca Candida albicans ATCC 10231

[a] Gram(+); [b] Gram(-); [c] - : Antimicrobial activity value is higher than 625 µg/mL

It is known that compounds containing benzimidazole moiety are known to play crucial roles in the structures and functions of a number of biologically important molecules. In addition, they have fluorescent characteristics. In this study, sixteen new benzimidazole derivatives (1–16) containing various substituted salicyl moieties were synthesized by cyclization of 4,5-dichloro/dimethyl-1,2-phenylenediamines with corresponding salicyl aldehydes. All target compounds were characterizated by using various physicochemical and spectroscopic techniques. It was found that the dichloro derivatives and 5-substitution on the phenolic ring redshift the fluorescence. Antimicrobial activities of the compounds were tested towards some selected Gram-positive and Gram- negative bacterial strains and C. albicans as fungi. It is seen that the dimethyl derivatives (9–16) show more antimicrobial activity towards the microorganisms than the dichloro derivatives. The antifungal effects of all the compounds against C. albicans in general are remarkable. 7 and 11 have superior antibacterial activity against S. epidermidis and S. aureus, respectively, with a MIC value of 2.4 µg/mL. Compound 13 shows the broadest activity, acting somewhat against all microorganisms.

As a conclusion, in this study, a new series of benzimidazole compounds were synthesized and their structural properties were examined, and the effects of different substituents on fluorescence properties and antimicrobial effects were examined, and remarkable findings were obtained.

All chemicals used in this work were supplied by Sigma-Aldrich, Merck and were used as any further purification. Melting points were recorded in glass capillary on a Buchi M-560 melting point apparatus. Elemental analysis data were determined with a Thermo Finnigan Flash EA 1112 analyzer. FT-IR spectra were recorded with a Bruker Optics Vertex 70 spectrometer. ¹H NMR spectra were determined in DMSO with a Varian Unity Inova 500 NMR spectrometer. Thermo Finnigan LCQ Advantage MAX LC/MS/MS was used to record Mass spectra. Fluorescence measurements were performed in EtOH (c = 1×10^− 4 mol/L) using a Shimadzu RF-5301 PC Spectrofluorophotometer. The compounds were prepared according to the literature procedure [25–26].

Synthesis of the compounds

3-(5,6-dichloro-1 H -benzimidazol-2-yl)benzene-1,2-diol (1): For synthesis of 1, 2,3-dihydroxybenzaldehyde (312 mg, 2.26 mmol) and an equivalent amount of NaHSO₃ (235 mg, 2.26 mmol) were reacted at room temperature in ethyl alcohol (10 cm³) for 4–5 hours. 4,5-dichlorobenzene-1,2-diamine (400 mg, 2.26 mmol) in DMF (10 cm³) was introduced to the reaction mixture and heated under reflux for 3 h (Scheme 2). After cooling to room temperature, the mixture was poured into cold water (100 cm³). The solid product which formed (1) was separated by filtration and re-crystallized from ethyl alcohol (573 mg, 86%). The compounds 2–8 were synthesized in a similar manner to 1. Beige powder. Mp: 317°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 13.30 (s, 1H, NH), 12.46 (s, 1H, OH2’), 9.27 (s, 1H, OH3’), 7.96 (s, 1H, H7), 7.92 (s, br, 1H, H4), 7.49 (dd, J = 7.8, 1.5 Hz, 1H, H6’), 6.94 (dd, J = 7.8, 1.5 Hz, 1H, H4’), 6.84 (t, J = 7.8, 7.8 Hz, 1H, H5’); FT-IR: ν˜ = 3546 m, 3285 m, 3058 w, 2974 w, 1665 m, 1640 m, 1594 m, 1536 m, 1496 m, 1332 m, 1270 s, 1239 s, 1095 m, 966 m, 863 m, 785 m, 665 m, 436 m ; fluorescence spectroscopy (ethanol, c = 1∙10–4 mol/L, nm): λ_max = 392 w, 498 m,br ; ESI-MS, m/z = 295.2 (100%, [M]+), 297.2 (63.4%, [M + 2]+); elemental analysis calcd. for C₁₃H₈Cl₂N₂O₂: C: 52.91, H: 2.73, N: 9.49; found: C: 52.88, H: 2.75, N: 9.48.

3-(5,6-dichloro-1 H -benzimidazol-2-yl)benzene-1,3-diol (2): 2 was synthesized in a similar manner to 1. 2,4-Dihydroxybenzaldehyde (312 mg, 2.26 mmol) was used. Yield: 607 mg, 91%. Brown powder. Mp: 305°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 13.02 (s,br, 1H, NH), 12.47 (s,br, 1H, OH2’), 10.07 (s, 1H, OH4’), 7.94 (s, 1H, H7), 7.86 (d, J = 8.8 Hz, 1H, H6’), 7.74 (s,br, 1H, H4), 6.45 (dd, J = 8.8, 2.4 Hz, 1H, H5’), 6.41 (d, J = 2.4 Hz, 1H, H3’); FT-IR: ν˜ = 3474 m, 3341 m, 3062 w, 2975 w, 1644 s, 1611 m, 1578 m, 1493 m, 1420 m, 1250 s, 1174 s, 1099 m, 971 m, 841 s, 794 s, 670 m, 486 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 404 m,br, 441 sh; elemental analysis calcd. for C₁₃H₈Cl₂N₂O₂: C: 55.81, H: 2.73, N: 9.49; found: C: 55.90, H: 2.71, N: 9.51.

3-(5,6-dichloro-1 H -benzimidazol-2-yl)benzene-1,4-diol (3): 3 was synthesized in a similar manner to 1. 2,5-Dihydroxybenzaldehyde (312 mg, 2.26 mmol) was used. Yield: 593 mg, 89%. Purple-grey powder. Mp: 355°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 13.10 (s,br, 1H, NH), 11.62 (s,br, 1H, OH2’), 9.12 (s, 1H, OH5’), 7.94 (s, 1H, H7), 7.88 (s,br, 1H, H4), 7.47 (d, J = 2.4 Hz, 1H, H6’), 6.87 (m, 2H, H3’+ H4’); FT-IR: ν˜ = 3555 m, 3312 m, 3100 w, 1656 m, 1621 m, 1598 m, 1528 s, 1496 m, 1451 s, 1260 s, 1201 m, 1165 s, 1091 m, 966 m, 859 m, 782 m, 657 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 397 m,br, 521 m,br; elemental analysis calcd. for C₁₃H₈Cl₂N₂O₂: C: 52.91, H: 2.73, N: 9.49; found: C: 55.94, H: 2.73, N: 9.52.

2-(5,6-dichloro-1 H -benzimidazol-2-yl)-4-methoxyphenol (4): 4 was synthesized in a similar manner to 1. 2-Hydroxy-5-methoxybenzaldehyde (334 mg, 2.26 mmol) was used. Yield: 537 mg, 77%. Mp: 358°C (decomp.); Reddish brown powder. ¹H NMR (DMSO-d₆, δ, ppm): 7.92 (s,br, 2H, H4 + H7), 7.66 (d, J = 2.9, 1H, H6’), 7.13 (dd, J = 8.8, 2.9 Hz, 1H, H4’), 6.98 (d, J = 8.8 Hz, 1H, H3’), 3.80 (s, 3H, OCH₃). FT-IR: ν˜ = 3301 m, 3060 w, 2968 w, 2840 w, 1631 m, 1608 m, 1591 w, 1537 m, 1449 m, 1374 m, 1251 s, 1097 m, 1057 s, 969 w, 858 m, 798 m, 733 m, 666 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 389 m,br, 511 m,br; elemental analysis calcd. for C₁₄H₁₀Cl₂N₂O₂: C: 54.51, H: 3.26, N: 8.91; found: C: 54.38, H: 3.29, N: 9.03.

2-(5,6-dichloro-1 H -benzimidazol-2-yl)-5-methoxyphenol (5): 5 was synthesized in a similar manner to 1. 2-Hydroxy-4-methoxybenzaldehyde (334 mg, 2.26 mmol) was used. Yield: 488 mg, 70%. Mp: 212°C (decomp.); dark yellow powder. ¹H NMR (DMSO-d₆, δ, ppm): 12.69 (s, 1H, NH), 8.23 (s, 1H, OH), 7.65 (d, J = 8.8 Hz, 1H, H6’), 7.38 (s, 1H, H7), 7.00 (s, 1H, H4), 6.64 (dd, J = 8.8, 2. 4 Hz, 1H, H5’), 6.56 (d, J = 2. 4 Hz, 1H, H3’), 3.87 (s, 3H, OCH₃); FT-IR: ν˜ = 3445 m, 3343 m, 3034 w, 2941 w, 2844 w, 1625 sh, 1608 s, 1517 m, 1487 m, 1440 m, 1391 m, 1287 m, 1120 s, 1028 m, 961 m, 889 m, 820 s, 797 m, 667m, 570 m, 478 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 w, 452 s; ESI-MS, m/z = 309.2 (100%, [M]⁺), 311.2 (67.1%, [M + 2]⁺); elemental analysis calcd. for C₁₄H₁₀Cl₂N₂O₂: C: 54.39, H: 3.26, N: 9.06; found: C: 54.34, H: 3.27, N: 9.02.

2-(5,6-dichloro-1 H -benzimidazol-2-yl)-6-methoxyphenol (6): 6 was synthesized in a similar manner to 1. 2-Hydroxy-3-methoxybenzaldehyde (334 mg, 2.26 mmol) was used. Yield: 593 mg, 85%. Mp: 341°C (decomp.); Light brown powder. ¹H NMR (DMSO-d₆, δ, ppm): 13.32 (s,br, 1H, NH), 12.48 (s, 1H, OH), 7.96 (s,br, 2H, H4 + H7), 7.63 (dd, J = 8.3, 1.5 Hz, 1H, H6’), 7.11 (dd, J = 8.3, 1.5 Hz, 1H, H4’), 6.96 (m, 1H, H5’), 3.84 (s, 3H, OCH₃); FT-IR: ν˜ = 3329 m, 3042 w, 2977 w, 2863 w, 1639 m, 1606 m, 1577 m, 1504 s, 1446 m, 1373 m, 1251 m, 1134 s, 1095 m, 970 m, 899 m, 837 m, 793 m, 684 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 w, 489 m, 512 sh; elemental analysis calcd. for C₁₄H₁₀Cl₂N₂O₂: C: 54.39, H: 3.26, N: 9.06; found: C: 54.39, H: 3.22, N: 9.05.

2-(5,6-dichloro-1 H -benzimidazol-2-yl)-4-methylphenol (7): 7 was synthesized in a similar manner to 1. 2-Hydroxy-5-methylbenzaldehyde (307 mg, 2.26 mmol) was used. Yield: 517 mg, 78%. Mp: 349°C (decomp.); Purple-grey powder. ¹H NMR (DMSO-d₆, δ, ppm): 13.21 (s,br, 1H, NH), 12.14 (s, 1H, OH), 7.98 (s,br, 1H, H7), 7.89 (d, J = 1.5 Hz, 1H, H6’), 7.82 (s,br, 1H, H4), 7.22 (dd, J = 8.3, 1.5 Hz, 1H, H4’), 6.95 (d, J = 8.3 Hz, 1H, H3’), 2.32 (s, 3H, CH₃); FT-IR: ν˜ = 3321 m, 3056 w, 2970 w, 2858 w, 1627 w, 1601 m, 1529 m, 1446 s, 1370 m, 1298 s, 1096 m, 1013 m, 971 m, 860 m, 787 s, 748 m, 665 m, 575 s, 490 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 393 w, 478 m,br; elemental analysis calcd. for C₁₄H₁₀Cl₂N₂O: C: 57.36, H: 3.44, N: 9.56: found; C: 57.38, H: 3.41, N: 9.53.

2-(5,6-dichloro-1 H -benzimidazol-2-yl)-6-methylphenol (8): 8 was synthesized in a similar manner to 1. 2-Hydroxy-3-methylbenzaldehyde (307 mg, 2.26 mmol) was used. Yield: 550 mg, 83%. Mp: 341°C (decomp.) ; Yellowish green powder. ¹H NMR (DMSO-d₆, δ, ppm): 13.49 (s,br 1H, NH), 12.99 (s, 1H, OH), 8.02 (s,br, 1H, H4), 7.86 (dd, J = 8.3, 1.0 Hz, 1H, H6’), 7.86 (s,br, 1H, H7), 7.31 (dd, J = 7.3, 1.0 Hz, 1H, H4’), 6.95 (t, J = 7.8, 7.3 Hz, 1H, H5’), 2.25 (s, 3H, CH₃); FT-IR: ν˜ = 3331 m, 3065 w, 2923 w, 1632 m, 1605 m, 1503 m, 1446 m, 1373 m, 1251 s, 1134 m, 1095 m, 871 m, 860 m, 794 s, 740 m, 661 m, 570 s, 421 m. ESI-MS, m/z = 293.3 (100%, [M]⁺), 295.3 (62.5%, [M + 2]⁺); fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 393 sh, 483 m,br; elemental analysis calcd. for C₁₄H₁₀Cl₂N₂O: C: 57.36, H: 3.44, N: 9.56; found: C: 57.35, H: 3.46, N: 9.52.

3-(5,6-dimethyl-1 H -benzimidazol-2-yl)benzene-1,2-diol (9): 9 was synthesized in a similar manner to 1. 2,3-Dihydroxybenzaldehyde (312 mg, 2.26 mmol) and 4,5-dimethyl-1,2-diamine (307 mg, 2.26 mmol) were used for synthesis. Yield: 431 mg, 75%. Cinnamon powder. Mp: 276°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 12.94 (s,br, 2H, NH + OH), 9.11 (s,br, 1H, OH), 7.50 (dd, J = 7.8, 1.5, H6’), 7.46 (s,br, 2H, H4 + H7), 6.93 (dd, J = 7.8, 1.5, H4’), 6.85 (t, J = 7.8, 7.8, H5’) 2.40 (s, 6H, 2×CH₃) ; FT-IR: ν˜ = 3573 m, 3293 m, 3050 w, 2970 m, 1615 m, 1601 m, 1580 m, 1475 s, 1335 m, 1270 s, 1100 m, 980 m, 813 m, 729 s, 599 m, 430 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 393 sh, 472 s; elemental analysis calcd. for C₁₅H₁₄N₂O₂: C: 70.85, H: 5.55, N: 11.02; found: C: 70.88, H: 5.53, N: 11.05.

3-(5,6-dimethyl-1 H -benzimidazol-2-yl)benzene-1,3-diol (10): 10 was synthesized in a similar manner to 9. 2,4-Dihydroxybenzaldehyde (312 mg, 2.26 mmol) was used. Yield: 464 mg, 81%. Yellowish green powder. Mp: 299°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 12.90 (s,br, 2H, NH + OH), 9.89 (s,br, 1H, OH_(4’)), 7.79 (d, J = 8.3 Hz, 1H, H6’), 7.35 (s, 2H, H4 + H7), 6.42 (dd, J = 8.3, 2.44 Hz, 1H, H5’), 6.37 (d, J = 2.44 Hz, 1H, H3’), 2.32 (s, 6H, 2×CH₃); FT-IR: ν˜ = 3626 m, 3275 m, 3050 w, 2977 w, 2940 m, 2924 m, 1623 m, 1608 s, 1578 w, 1491 m, 1466 m, 1392 m, 1311 m, 1235 m, 1145 s, 977 m, 860 m, 799 m, 664 m, 524 m, 476 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 379 sh, 426 s; elemental analysis calcd. for C₁₅H₁₄N₂O₂: C: 70.85, H: 5.55, N: 11.02; found: C: 70.83, H: 5.57, N: 11.05.

3-(5,6-dimethyl-1 H -benzimidazol-2-yl)benzene-1,4-diol (11): 11 was synthesized in a similar manner to 9. 2,5-Dihydroxybenzaldehyde (312 mg, 2.26 mmol) was used. Yield: 402 mg, 70%. Milky brown powder. Mp: 324°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 9.22 (s, 1H, OH_(5’)), 7.49 (s, 2H, H4 + H7), 7.39 (d, J = 2.4 Hz, 1H, H6’), 6.93 (s, 2H, H3’+H4’ ), 2.36 (s, 3H, CH₃₍₅₎), 2.27 (s, 3H, CH₃₍₆₎); FT-IR: ν˜ = 3382 m, 3192 m, 3047 w, 2921 w, 1633 m, 1618 m, 1553 m, 1470 m, 1382 m, 1225 m, 1094 s, 987 m, 858 m, 743 m, 683 m, 591 m, 472 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 393 sh, 495 m; elemental analysis calcd. for C₁₅H₁₄N₂O₂: C: 70.85, H: 5.55, N: 11.02; found: C: 70.85, H: 5.49, N: 11.03.

2-(5,6-dimethyl-1 H -benzimidazol-2-yl)-4-methoxyphenol (12): 12 was synthesized in a similar manner to 9. 2-Hydroxy-5-methoxybenzaldehyde (344 mg, 2.26 mmol) was used. Yield: 455 mg, 76%. Milky brown powder. Mp: 293°C (decomp.); ¹H NMR (DMSO-d₆, δ, ppm): 7.61 (d, J = 2.9 Hz, 1H, H6’), 7.43 (s, 1H, H4), 6.99 (s, 1H, H7), 6.97 (t, J = 8.8, 2.9 Hz, 1H, H4’), 6.75 (d, J = 8.8 Hz, 1H, H3’), 3.80 (s, 3H, OCH₃), 2.35 (s, 3H, 2×CH₃) ; FT-IR: ν˜ = 3543 m, 3320 m, 3017 w, 2971 m, 2857 w, 1623 w, 1600 m, 1592 w, 1478 m, 1382 m, 1251 s, 1105 m, 1062 s, 998 m, 851 m, 782 m, 732 m, 603 m, 566 m, 433 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 sh, 460 m,br; elemental analysis calcd. for C₁₆H₁₆N₂O₂: C: 71.62, H: 6.01, N: 10.44; found: C: 71.65, H: 6.02, N: 10.45.

2-(5,6-dimethyl-1 H -benzimidazol-2-yl)-5-methoxyphenol (13): 13 was synthesized in a similar manner to 9. 2-Hydroxy-4-methoxybenzaldehyde (344 mg, 2.26 mmol) was used. Yield: 490 mg, 81%. Mp: 281°C (decomp.); Yellowish green powder. ¹H NMR (DMSO-d₆, δ, ppm): 7.90 (d, 1H, J = 8.8 Hz, H6’), 7.38 (s,br, 1H, H4), 7.36 (s,br, 1H, H7), 6.60 (dd, 1H, J = 8.8, 2.4 Hz, H5’), 6.56 (d, 1H, J = 2.4, H3’), 3.80 (s, 3H, OCH₃), 2.33 (s, 6H, 2×CH₃); FT-IR: ν˜ = 3353 m, 3062 w, 2965 m, 2841 w, 1660 s, 1616 m, 1452 m, 1380 m, 1289 s, 1169 m, 1037 s, 841 s, 756 m, 679 m, 571 m, 439 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 381 sh, 422 s; elemental analysis calcd. for C₁₆H₁₆N₂O₂: C: 71.62, H: 6.01, N: 10.44; found: C: 71.61, H: 6.04, N: 10.40.

2-(5,6-dimethyl-1 H -benzimidazol-2-yl)-6-methoxyphenol (14): 13 was synthesized in a similar manner to 9. 2-Hydroxy-3-methoxybenzaldehyde (344 mg, 2.26 mmol) was used. Yield: 454 mg, 75%. Mp: 293°C (decomp.); Light yellow powder. ¹H NMR (DMSO-d₆, δ, ppm): 13.36 (s, 1H, NH), 12.91 (s, 1H, OH), 7.59 (dd, 1H, J = 7.8, 1.5 Hz, H6’), 7.49 (s, 1H, H4), 7.35 (s, 1H, H7), 7.05 (dd, 1H, J = 8.3, 1.5 Hz, H5’), 6.92 (t, 1H, J = 8.3, 7.8 Hz, H4’), 3.82 (s, 3H, OCH₃), 2.35 (s, 3H, CH₃₍₆₎), 2.33 (s, 3H, CH₃₍₅₎); FT-IR: ν˜ = 3329 m, 3253 m, 3051 w, 2978 m, 2829 w, 1624 m, 1599 m, 1504 s, 1461 m, 1384 m, 1222 s, 1169 m, 1048 m, 852 m, 760 m, 685 w, 578 m, 470 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 w, 492 m,br; elemental analysis calcd. for C₁₆H₁₆N₂O₂: C: 71.62, H: 6.01, N: 10.44; found: C: 71.60, H: 6.01, N: 10.42.

2-(5,6-dimethyl-1 H -benzimidazol-2-yl)-4-methylphenol (15): 15 was synthesized in a similar manner to 9. 2-Hydroxy-5-methylbenzaldehyde (307 mg, 2.26 mmol) was used. Yield: 450 mg, 79%. Mp: 285°C (decomp.); Light yellow powder. ¹H NMR (DMSO-d₆, δ, ppm): 12.90 (s, 1H, NH), 12.81 (s, 1H, OH), 7.82 (d, J = 1.2 Hz, 1H, H6’), 7.49 (s,br, 1H, H4), 7.46 (s,br, 1H, H7), 7.15 (dd, J = 8.3, 1.5 Hz, 1H, H4’), 6.90 (dd, J = 8.3, 1.8 Hz, 1H, H5’), 2.34 (s, 3H, CH₃₍₆₎), 2.33 (s, 3H, CH₃₍₅₎), 2.31 (s, CH_3(3’)); FT-IR: ν˜ = 3340 s, 3030 m, 2969 m, 2858 w, 1636 m, 1603 m, 1532 s, 1472 m, 1379 m, 1254 s, 1165 m, 1014 m, 848 m, 786 s, 629 m, 560 m, 436 m; fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 w, 460 m; elemental analysis calcd. for C₁₆H₁₆N₂O: C: 76.16, H: 6.39, N: 11.10; found: C: 76.15, H: 6.41, N: 11.10.

2-(5,6-dimethyl-1 H -benzimidazol-2-yl)-6-methylphenol (16): 16 was synthesized in a similar manner to 9. 2-Hydroxy-3-methylbenzaldehyde (307 mg, 2.26 mmol) was used. Yield: 410 mg, 72%. Mp: 314°C (decomp.); Milky brown powder. ¹H NMR (DMSO-d₆, δ, ppm): 13.55 (s, 1H, NH), 12.91 (s, 1H, OH), 7.82 (dd, J = 7.8, 1.5 Hz, 1H, H6’), 7.47 (s,br, 1H, H4), 7.34 (s,br, 1H, H7), 7.23 (d, J = 7.3 Hz, 1H, H4’), 6.89 (t, J = 7.8, 7.3 Hz, 1H, H3’), 2.35 (s, 3H, CH₃₍₆₎), 2.33 (s, 3H, CH₃₍₅₎), 2.25 (s, CH_3(5’)); FT-IR: ν˜ = 3268 m, 3048 m, 2975 m, 2862 w, 1640 m, 1604 m, 1505 s, 1439 m, 1383 m, 1255 s, 1164 m, 1003 m, 872 m, 804 s, 741 m, 664 m, 552 m, 436 m. ESI-MS, m/z = 253.3 (100%, [M + 1]⁺), 254.3 (20.0%, [M + 2]⁺); fluorescence spectroscopy (ethanol, c = 1∙10^− 4 mol/L, nm): λ_max = 394 w, 444 m,br; elemental analysis calcd. for C₁₆H₁₆N₂O: C: 76.16, H: 6.39, N: 11.10; found: C: 76.13, H: 6.37, N: 11.12.

Determination of antimicrobial activity

Antimicrobial activity of the obtained compounds (1–16) were evaluated by using the microbroth dilutions method, according to guidelines outlined by the Clinical Laboratory Standards Institute (CLSI) [27–28]. The bacteria including S. aureus (ATCC 6538), S. epidermidis (ATCC 12228), E. coli (ATCC 8739), K. pneumoniae (ATCC 4352), P. aeruginosa (ATCC 27853), P. mirabilis (ATCC 14153) and C. albicans (ATCC 10231) were used for antimicrobial screening. Mueller-Hinton broth (MHB) and RPMI-1640, buffered with MOPS at pH = 7.0 were used as a broth media for bacteria and yeast, respectively. All test compounds were dissolved in DMSO to give stock solutions. 2-fold serial dilutions were prepared in the medium having concentration in a range of 4.9 to 5000 µg/mL. The inoculum was prepared from a 14–18 h broth culture of the tested bacteria and 24 h culture of yeast strains. Then, the suspensions were adjusted to match the turbidity of a 0.5 McFarland standard and further diluted in the corresponding culture medium to obtain the required inoculum concentration of 5x10⁵ CFU/mL for bacteria and 0.5x10³ to 2.5x10³ CFU/mL for yeast in the test plates. To prevent evaporations, the plates were covered with plastic bags. The plates were incubated at 35 ºC for about 18 to 20 h and 46 to 50 h for antibacterial and antifungal activity. After incubation, the minimum inhibitory concentrations (MIC) were defined as the lowest concentration of compounds giving complete inhibition of visible growth. Ciprofloxacin for bacteria and fluconazole for fungi were used as positive controls whereas DMSO was used as a negative control. Based on values of these controls, the results were obtained. Each experiments were repeated three times in order to minimize the error.

Acknowledgement

This work was financially supported by Scientific Research Projects Coordination Unit of Istanbul University-Cerrahpaşa.

Ethical Approval

No ethical approval required.

Competing interests

No competing interests.

Authors' contributions

Demet Gürbüz synthesized the compounds and wrote the main manuscript text.

Aydin Tavman and Adem Çınarlı characterized the compounds.

A. Seher Birteksöz Tan evaluated the antimicrobial activity tests.

Funding

Scientific Research Projects Coordination Unit of Istanbul University-Cerrahpaşa.

Availability of data and materials

The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

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Scheme 1 and 2 are available in Supplementary Files section.

No competing interests reported.

Scheme1.jpg
Scheme 1 Structure of the compounds in the study
Scheme2.jpg
Scheme 2 Synthesis mechanism for the compounds 1 – 16
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Synthesis, Characterization and Antimicrobial Activity of Some New Benzimidazole Derivatives Bearing the Salicyl Moiety

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Abstract

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Introduction

Result And Discussion

FT-IR Spectra

Fluorescence Spectra

Antimicrobial Activity

Conclusion

Experimental Section

Synthesis of the compounds

Determination of antimicrobial activity

Declarations

References

Scheme

Competing Interests

Supplementary Files

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